School of Life Sciences, The Biodesign Institute, P.O. Box 875401, Arizona State University, Tempe, AZ 85287-5401, USA.
J Virol. 2010 Nov;84(21):11418-28. doi: 10.1128/JVI.01131-10. Epub 2010 Aug 18.
Coronavirus membrane (M) proteins play key roles in virus assembly, through M-M, M-spike (S), and M-nucleocapsid (N) protein interactions. The M carboxy-terminal endodomain contains a conserved domain (CD) following the third transmembrane (TM) domain. The importance of the CD (SWWSFNPETNNL) in mouse hepatitis virus was investigated with a panel of mutant proteins, using genetic analysis and transient-expression assays. A charge reversal for negatively charged E(121) was not tolerated. Lysine (K) and arginine (R) substitutions were replaced in recovered viruses by neutrally charged glutamine (Q) and leucine (L), respectively, after only one passage. E121Q and E121L M proteins were capable of forming virus-like particles (VLPs) when coexpressed with E, whereas E121R and E121K proteins were not. Alanine substitutions for the first four or the last four residues resulted in viruses with significantly crippled phenotypes and proteins that failed to assemble VLPs or to be rescued into the envelope. All recovered viruses with alanine substitutions in place of SWWS residues had second-site, partially compensating, changes in the first TM of M. Alanine substitution for proline had little impact on the virus. N protein coexpression with some M mutants increased VLP production. The results overall suggest that the CD is important for formation of the viral envelope by helping mediate fundamental M-M interactions and that the presence of the N protein may help stabilize M complexes during virus assembly.
冠状病毒膜（M）蛋白在病毒组装中发挥关键作用，通过 M-M、M-刺突（S）和 M-核衣壳（N）蛋白相互作用。M 羧基末端内结构域在第三个跨膜（TM）结构域之后包含一个保守结构域（CD）。使用遗传分析和瞬时表达测定，用一组突变蛋白研究了小鼠肝炎病毒中 CD（SWWSFNPETNNL）的重要性。带负电荷的 E(121) 的电荷反转不能被容忍。赖氨酸（K）和精氨酸（R）取代在仅传代一次后，分别被带中性电荷的谷氨酰胺（Q）和亮氨酸（L）取代，从而在恢复的病毒中取代。当与 E 共表达时，E121Q 和 E121L M 蛋白能够形成病毒样颗粒（VLPs），而 E121R 和 E121K 蛋白则不能。前四个或后四个残基的丙氨酸取代导致病毒表型严重受损，且蛋白无法组装 VLPs 或无法拯救到包膜中。SWWS 残基被丙氨酸取代的所有恢复病毒都具有第二位置、部分补偿的 M 的第一个 TM 变化。脯氨酸的丙氨酸取代对病毒几乎没有影响。N 蛋白与一些 M 突变体的共表达增加了 VLP 的产生。总体结果表明，CD 对于形成病毒包膜很重要，通过帮助介导基本的 M-M 相互作用，并且 N 蛋白的存在可能有助于在病毒组装过程中稳定 M 复合物。
